Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.26.9 (ribonuclease)
6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The estrogen receptor (ER) acts as a transcription factor to regulate multiple cellular functions involved in normal physiology, differentiation, and reproduction. To date, there is no known animal model for studying aberrant ER expression. Therefore, we created transgenic mice expressing the wild-type mouse ER under the control of the mouse metallothionein-I (MT) promoter to determine whether overexpression of the ER would disrupt normal reproductive processes. Five male and one female founder mice were produced, and all were fertile. The progeny from these mice were screened for MT-mER expression by the ribonuclease protection assay. Mice in all six lines were found to express the transgene in a variety of tissues, although generally at low levels. The highest level of expression was observed in the female reproductive tract of line E. Females in all six lines demonstrated aberrant reproductive phenotypes involving processes at parturition and, with some of the lines, a tendency toward reduced fertility. Gestational length was prolonged up to 4 days beyond the normal gestation of 19 days, providing evidence of delayed parturition. In addition, prolonged labor (up to 3 days in length to deliver all pups) and labors requiring cesarean sections for maternal survival demonstrated the occurrence of dystocia in the MT-mER females. As maternal age increased, the incidence of stillborn litters, delayed parturition, and dystocia approached 100% in the transgenic dams. Difficulties at parturition were not observed in nontransgenic control females. These phenotypes suggest that the mechanisms regulating parturition may be perturbed by improper expression of the ER. The MT-mER transgenic mice may provide a novel approach for studying the estrogen-regulated signals involved in parturition and fertility as well as a unique animal model for the human reproductive phenotypes of delayed parturition and dystocia.
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PMID:Aberrant reproductive phenotypes evident in transgenic mice expressing the wild-type mouse estrogen receptor. 801 72

Recent evidence suggests that the expression of estrogen receptor (ER) variants in breast cancer may interfere with wild-type (wt) ER function and be related to tumor progression and resistance to hormone treatment. One of these variants, ER delta E5, lacking that part of the hormone-binding domain encoded by exon 5, has previously been identified in breast tumors with the unusual estrogen receptor negative (ER-) and progesterone receptor positive (PgR+) phenotype and found to possess constitutive and hormone-independent transcriptional activity. Using a ribonuclease protection assay, we analyzed 27 breast tumors and 4 breast cell lines for the presence of this variant. We found the ER delta E5 variant to be expressed, not only in all of three ER-/PgR+ tumors but also in 19 of 20 ER+/PgR+ or ER+/PgR- tumors. Moreover, the variant was always coexpressed with and often in excess of wtER. ER delta E5 was also found in three breast cancer cell lines (MCF7, T47D, and ZR75-1), although to a lesser extent than wtER. A complete absence of both ER delta E5 and wtER was noted in four ER-/PgR- tumors and one normal breast cell line (HBL-100). Thus, our data suggest that the occurrence of ER delta E5 in breast cancer may represent a critical stage in tumor progression to autonomy.
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PMID:An exon 5 deletion variant of the estrogen receptor frequently coexpressed with wild-type estrogen receptor in human breast cancer. 826 97

Mechanisms regulating responses of the ovine uterus to endocrine and paracrine signals during the estrous cycle and pregnancy are likely to require tissue- and cell-specific regulation of steroid hormone receptor gene expression. To determine effects of day and pregnancy status (cyclic or pregnant) on uterine estrogen receptor (ER) and progesterone receptor (PR) gene expression, ewes were hysterectomized either on Day 1 (Day 0 = estrus/mating), 6, 11, 13, or 15 of the estrous cycle (n = 3/day) or on Day 11, 13, 15, 17, or 25 of early pregnancy (n = 5/day). Steady state levels of ER and PR mRNA were determined in endometrial and myometrial tissues by slot-blot hybridization and ribonuclease protection assays, respectively, using homologous ovine ER and PR cRNA probes. Changes in spatial expression of ER and PR mRNA and protein in uterine tissue sections were determined by in situ hybridization and immunocytochemical analyses. In cyclic ewes, steady state levels of endometrial ER mRNA were highest on Day 1, declined between Days 1 and 6, and increased between Days 11 and 15. However in pregnant ewes, endometrial ER mRNA levels decreased between Days 11 and 15 and increased slightly between Days 15 and 25. In cyclic ewes, levels of myometrial ER mRNA were highest on Day 1, decreased to Day 6, and remained low thereafter. In cyclic ewes, endometrial PR mRNA levels were highest on Day 1, decreased between Days 1 and 11, and then increased between Days 13 and 15. In cyclic ewes, myometrial PR mRNA levels were highest on Day 1 and declined thereafter. Endometrial PR mRNA levels were not different between cyclic and pregnant ewes on Days 11, 13, and 15. In pregnant ewes, PR mRNA levels were low on Day 11, increased between Days 11 and 17, and decreased between Days 17 and 25. In pregnant ewes, myometrial PR mRNA levels were low and did not change between Days 11 and 25. In situ hybridization and immunocytochemical analyses revealed distinct tissue- and cell type-specific alterations in uterine ER and PR mRNA and protein expression during the estrous cycle and early pregnancy that generally paralleled overall changes in steady state levels of ER and PR mRNAs. In the endometrium, the most striking observation was that PR mRNA and protein expression disappeared from the luminal and shallow glandular epithelium between Days 6 and 13 of the estrous cycle, whereas ER mRNA and protein expression was low on Days 6 and 11 and increased between Days 11 and 15 in the luminal and shallow glandular epithelium. During early pregnancy, expression of ER and PR mRNAs, as well as ER and PR protein, was very low or absent in the luminal and shallow glandular epithelium between Days 13 and 25 of pregnancy. Moreover, ER and PR mRNA and protein were consistently present at low levels in the stroma and deep glandular epithelium in both cyclic (Days 11-15) and pregnant (Days 11-25) ewes. Collectively, results suggest that uterine ER and PR gene expression is regulated in a tissue- and cell type-specific manner during the estrous cycle and early pregnancy.
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PMID:Temporal and spatial alterations in uterine estrogen receptor and progesterone receptor gene expression during the estrous cycle and early pregnancy in the ewe. 856 11

Several studies in the past few years have supported the hypothesis that oxytocin (OT) is synthesized in a paracrine system within the pregnant human uterus and that this paracrine system may be an important regulator of the timing of human parturition. Using ribonuclease protection assays, we have demonstrated a three-fold increase in the rate of synthesis of OT mRNA in human decidua around the time of parturition. We also have shown that a similar increase in OT mRNA and peptide synthesis can be stimulated in vitro by physiological concentrations of estradiol. This increase is inhibited by concomitant use of the estrogen receptor (ER) blocker tamoxifen or by transcription inhibitors. Progesterone had little, if any effect. We also detected mRNAs for ER and progesterone receptor (PR) in amnion, chorion and decidua with the same relative tissue concentrations as OT mRNA. The concentrations of ER but not PR increased significantly around the time of labour onset. To determine if local OT concentrations may be regulated by changes in OT metabolism, we determined kinetic parameters for OT metabolism in decidua, chorion and placenta. [3H]tyrosyl-OT was used as substrate. Metabolites were separated using HPLC and identified using amino acid analysis and mass spectrometry. Metabolism in decidua and chorion occurred predominantly via a cytosolic post-proline endopeptidase and the activity was comparable to placenta. In microsomal fractions, cystine aminopeptidase activity predominated and placenta had significantly more activity than decidua and chorion. There were no changes in any Km or apparent vmax values around the time of parturition. These findings support the existence of a paracrine system within human decidua that involves sex steroids regulating synthesis of OT and that undergoes significant changes around the time of parturition. Changes in local OT concentrations are controlled by rates of synthesis rather than rates of metabolism.
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PMID:Synthesis and metabolism of oxytocin in late gestation in human decidua. 871 92

The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) induces severe reproductive defects in male rats when exposure occurs in utero and during lactation. Yet there is currently a paucity of information regarding the effects of this exposure paradigm in females. In the current study, we examine the effects of TCDD during fetal and perinatal development on the estrogen-signaling system in peripubertal female rats. Pregnant Holtzman rats were given 1 microgram/kg TCDD or vehicle control by gavage on gestational Day 15. Body weights were reduced, though not significantly, on postnatal Day 21. While ovarian and uterine wet weights were not increased by TCDD exposure, the percentage of body weight attributed to the ovary was increased significantly. Through use of ribonuclease protection and gel-shift assays, exposed females were compared with nonexposed counterparts for estrogen receptor (ER) mRNA and DNA-binding activity in the following tissues: hypothalamus, pituitary (mRNA only), uterus, and ovary. ER mRNA levels increased in the hypothalamus, uterus, and ovary, and decreased in the pituitary. The results of the DNA-binding assays paralleled the mRNA results in the uterus, while DNA-binding activity was decreased in the hypothalamus and was unchanged in ovarian protein extracts. Circulating concentrations of estrogen were significantly lower in TCDD-exposed rats than in controls. These data suggest that the decrease in serum estrogen may be a cause of the alterations in ER mRNA; the changes in ER DNA-binding activity may indicate alterations in either translation or posttranslational receptor processing. Overall, this study shows that TCDD may act systemically in this model, and these effects should not necessarily be characterized as antiestrogenic.
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PMID:In utero and lactational exposure of female Holtzman rats to 2,3,7,8-tetrachlorodibenzo-p-dioxin: modulation of the estrogen signal. 879 59

Studies have shown that the pineal gland via its hormone, melatonin, induces the involution of male and female reproductive systems in seasonally reproducing animals. Melatonin has direct inhibitory effects on both hypothalamic and pituitary functions, which are also exquisitely sensitive to the feedback effects of estradiol. Since melatonin can modulate estrogen receptor (ER) expression in other tissues, immunocytochemical and ribonuclease protection analyses were used to examine the effects of 12 weeks of daily late afternoon injections of melatonin on ER protein and mRNA levels in the hypothalamus of Lak.LVG golden hamsters. Significant decreases in ER-immunoreactivity were noted in the medial preoptic area (MPOA) and bed nucleus of the stria terminalis (BNST) in response to melatonin, while other hypothalamic areas which express ER, e.g. the anterior hypothalamus, showed less dramatic changes. Hypothalamic ER mRNA was decreased in response to melatonin in both intact and ovariectomized animals by 25%. In intact, cycling female hamsters, there was a significant reduction in uterine weight after melatonin treatment. These results suggest that melatonin exerts its anti-reproductive effects in hamsters by modulating ER levels in neurons of the MPOA and BNST, thereby influencing steroid feedback mechanisms.
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PMID:Effects of melatonin on estrogen receptor expression in the forebrain of outbred (Lak.LVG) golden hamsters. 911 84

Oxytocin (OT) and its receptor (OTR) are synthesized in the endometrium and myometrium of the pregnant rat during late gestation. Both are regulated by estrogen and progesterone (P4), and tissue concentrations of both increase markedly before parturition. The P4 antagonist RU486 will induce parturition in the rat. The purpose of the present studies was to investigate changes in OT and OTR messenger RNA (mRNA) and peptide synthesis within the pregnant rat uterus during RU486-induced parturition. Pregnant rats were given a single injection of RU486 (2.5 mg/rat in oil) on day 15 of pregnancy (normal delivery occurs on day 22). Control animals received injections of oil only. Groups of animals (n = 5 in each group) were euthanized at 0, 6, 12, 24, and 48 h after injection and during labor (immediately after delivery of the first pup). Maternal serum estradiol (E2), P4 and uterine OT, and PGE2 concentrations were measured by RIA. Prostaglandin F2alpha and estrogen receptor levels were measured by enzyme immunoassay (EIA). OTR and P4 receptor (PR) were measured using radioligand-binding assays. OT, OTR, and estrogen receptor mRNAs were measured with ribonuclease protection assays. The average time to delivery, after RU486 injection, was 27.0 +/- 1.2 h. Serum E2 and P4 levels were increased slightly, but significantly, at 24 h after RU486. In controls, OT mRNA increased significantly, and this increase was blocked in the RU486 treatment group. OTR mRNA levels increased within 6 h of RU486 and remained elevated until delivery. OTR peptide was increased by 12 h. PGE2 and PGF2alpha were increased 3-fold and 16-fold, respectively, but not until after the increase in OTR had occurred. We conclude that the mechanism of action of RU486 is to inhibit the P4 suppression of OTR synthesis, allowing increased expression of OTR, which may directly stimulate myometrial contractions or act indirectly through increased synthesis of PGs.
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PMID:Effects of RU486 on estrogen, progesterone, oxytocin, and their receptors in the rat uterus during late gestation. 920 15

The conventional methods for mRNA quantitation such as Northern blotting or ribonuclease protection assay sometimes lack enough sensitivity to study low abundance mRNAs or to work with limited amounts of biological samples. The sensitivity of the polymerase chain reaction (PCR) linked to reverse transcription (RT-PCR) has proven useful in amplifying specific mRNAs, especially those present in low copy number. Though, the quantitation of nucleic acids by means of PCR has proven problematic. The main constraint in obtaining quantitative data is inherent in the amplification reaction. Because amplification is an exponential process, small variations in the efficiency of amplification may significantly affect the final yield of the PCR product. The variables that influence the rate of the PCR include the abundance of the mRNA present in the starting material, the concentrations of the Taq DNA polymerase, dNTPs and magnesium ions, the annealing and elongation conditions, the ramping temperatures and the formation of primer secondary structures. Moreover, with the progression of the PCR cycles, reagents are consumed and inhibitors generated, leading to non-linear synthesis of DNA. Finally, tube-to-tube variations sometimes preclude accurate quantitation. Most of the above-mentioned problems can be overcome by the choice of adequate internal controls. The present report reviews two recently developed methods for RNA quantitation, the semi-quantitative PCR and the quantitative PCR illustrated for the measurement of monoamine oxidase (MAO) A and B mRNAs and the estrogen receptor (ER) mRNA respectively, with a particular emphasis on the design of appropriate internal controls to compensate for the intra- and inter-assay variability inherent to RT-PCR.
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PMID:Quantitation of low abundance mRNAs in glial cells using different polymerase chain reaction (PCR)-based methods. 938 56

Previous experiments have demonstrated that C-type natriuretic peptide (CNP) expression in the uterus varies during the estrous cycle with maximal expression at proestrus. The present study was designed to determine whether exogenous steroid hormones regulate uterine CNP expression in ovariectomized mice. Estradiol increased significantly (3-fold) uterine immunoreactive CNP (irCNP) rapidly and dose dependently in ovariectomized mice as measured by radioimmunoassay. Other steroids produced either no significant change (testosterone, 1 mg; 2-methoxyestradiol, 1 microgram) or weak induction (estriol, 1 microgram) from vehicle controls. Progesterone (1 mg) significantly attenuated the estrogen-stimulated irCNP response by 50% when injected 30 min before estrogen (1 microgram) in estrogen-primed ovariectomized mice. Estrogen-stimulated increases in uterine CNP transcripts detected by ribonuclease protection analyses were blocked by actinomycin D (160 micrograms) or ICI-164,384 (20 micrograms), a specific nuclear estrogen receptor antagonist. These results indicate that a nuclear estrogen receptor is required for estrogen to stimulate uterine CNP transcription and that progesterone negatively regulates estrogen-stimulated CNP expression.
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PMID:Estradiol induces C-type natriuretic peptide gene expression in mouse uterus. 943 3

The effects of estradiol treatment, which stimulates cell division in rat uterine epithelial cells, on the in vivo expression of heparin-binding epidermal growth factor (HB-EGF), cyclin D1, and cyclin B1 messenger RNA (mRNA) in these cells have been examined using ribonuclease protection assays. Estradiol gave rise to significant increases in steady state levels of HB-EGF 2 and 24 h after treatment. Cyclin D1 mRNA levels were elevated 8 and 10 h after estradiol administration, corresponding to the G1 phase of the mitotic cycle, and cyclin B1 mRNA was only expressed 16-24 h after estradiol treatment, which corresponds to the G2 and M phases of the rat uterine epithelial cell cycle. Estradiol-stimulated increases in HB-EGF mRNA were not affected by treatment with cycloheximide, but were inhibited by the estrogen antagonist compound, ICI 164,384, demonstrating that the estrogen-stimulated increase in HB-EGF mRNA is a primary, estrogen receptor-mediated response of rat uterine epithelium to estradiol. Progesterone treatment, which blocks epithelial cells in G1 of the cycle, suppressed levels of HB-EGF mRNA below those observed in ovariectomized rats. These results indicate that HB-EGF mediates the regulatory effects of both estradiol and progesterone on rat uterine epithelial cell proliferation through an effect on the production of G1 phase molecules such as cyclin D1.
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PMID:Mediators of estradiol-stimulated mitosis in the rat uterine luminal epithelium. 949 26


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